Malate dehydrogenase



Malate Dehydrogenase (MDH)(PDB entry 2x0i) is most known for its role in the metabolic pathway of the tricarboxylic acid cycle, also know as the Kreb's Cycle, which is critical to cellular respiration in cells ; however, the enzyme is also in many other metabolic pathways such as glyoxylate bypass, amino acid synthesis, glucogenesis, and oxidation/reduction balance. It is classified as an oxidoreductase. Malate Dehydrogenase has been extensively studied due to its many isozymes. The enzyme exists in two places inside a cell: the mitochondria and cytoplasm. In the mitochondria, the enzyme catalyzes the reaction of malate to oxaloacetate; however, in the cytoplasm, the enzyme catalyzes oxaloacetate to malate to allow transport. This conversion is an essential catalytic step in each different metabolic mechanism. The enzyme malate dehydrogenase is composed of either a dimer or tetramer depending on the location of the enzyme and the organism it is located in. During catalysis, the enzyme subunits are non-cooperative between active sites. The mitochondrial MDH is complexly, allosterically controlled by citrate, but no other known metabolic regulation mechanisms have been discovered. Further, the exact mechanism of regulation has yet to be discovered. Kinetically, the pH of optimization is 7.6 for oxaloacetate conversion and 9.6 for malate conversion. The reported K(m) value for malate conversion is 215 uM and the V(max) value is 87.8 uM/min.



Structure
The secondary structure of a single subunit contains a 9 beta sheet parallel backbone wrapped by 9 large alpha helices. Near the sodium bound end, 4 small anti-parallel beta sheets and 1 small alpha helix enable a turn in the residue chain(small turn). Opposite the sodium bound ligand, 6 alpha helices point towards a common point, three on each side of the beta sheet backbone. The alpha helices form a small groove for a NAD+ cofactor to attach near the beta sheeting. The structure most nearly resembles an alternating alpha/beta classification. As for the 3D structure, the enzyme forms a sort of crevice for the substrate to bind.

Mechanism
The mechanism of catalysis is dependent on several invariant residues. These residues are HIS 195 and ASP 168, which are both involved in hydrogen bonding, ASP 53 associated with NAD+ binding, and a triad of arginine residues at 102, 109, and 171. During the conversion of malate to oxaloacetate, a key conformational change occurs on the binding of substrate in which a “loop” flips into an up position to block the active site from the solvent. When this occurs, the other residues in the active site are brought closer to the substrate to enable the conversion. R102 and R109 are involved in this loop flip and thus invariant. After the loop flip, the malate complex is stabilized via hydrogen bonding before accepting a proton transfer from NADH to form oxaloacetate.

Evolutionary Divergence
The evolutionary past of MDH shows a divergence to form lactate dehydrogenase (LDH) which functions in a very similar way to MDH. Although there is a very low sequence conservation among MDH and LDH’s the structure of the enzyme has remained relatively conserved. The key difference between the two is in the substrate: LDH catalyzes pyruvate to lactate.

3D Structures of Malate Dehydrogenase
The holo-MDH contains NAD or its derivatives while the apo-MDH lacks it.

Holo-MDH
2x0r – HmMDH (mutant)+NAD - Haloarcula marismortui

1o6z - HmMDH (mutant)+NADH

1hlp – HmMDH+NAD

1x0i – AfMDH+NADH – Archaeoglobus fulgidus

2x0j - AfMDH+etheno-NAD

1hlp – HmMDH+NAD

1x0i – AfMDH+NADH

2x0j - AfMDH+etheno-NAD

1ib6, 1ie3 – EcMDH (mutant)+NAD - Escherichia coli

1emd – EcMDH+NAD+citrate

3i0p – MDH+NAD – Entamoeba histolytica

3gvh – BmMDH+NAD – Brucella melitensis

3gvi - BmMDH+ADP

2hjr – MDH+adenosine diphosphoribose – Cryptosporidium parvum

2dfd – MDH+NAD – human type 2

1wze – TfMDH (mutant)+NAD – Thermus flavus

1wzi - TfMDH (mutant)+NDP

1bdm - TfMDH (mutant)+beta-6-hydroxy-1,4,5,6-tetrhydronicotinamide adenine dinucleotide

1bmd – TfMDH+NAD

1y7t – TtMDH+NADPH – Thermus thermophilus

2cvq - TtMDH+NADP

1v9n – MDH+NADPH – Pyrococcus horikoshii

1z2i – MDH+NAD – Agrobacterium tumefaciens

1uxg, 1uxh, 1uxi, 1uxj, 1uxk, 1ur5 – MDH (mutant)+NAD – Chloroflexus aurantiacus

1guz, 1guy, 1gv0 – CvMDH+NAD – Chlorobium vibrioforme

1civ – MDH+NADP – Flaveria bidentis

1b8u, 1b8v – AaMDH+NAD - Aquaspirillum arcticum

5mdh – SsMDH+NAD+alpha-ketomalonic acid – Sus scrofa

4mdh – SsMDH+NAD

apo-MDH
2j5r, 2j5k, 2j5q, 1d3a – HmMDH

2hlp – HmMDH (mutant)

3hhp, 2pwz – EcMDH

3fi9 – MDH – Porphyromonas gingivalis

3d5t - MDH – Burkholderia pseudomallei

2d4a – MDH – Aeropyrum pernix

1iz9 - TtMDH

1sev, 1smk – MDH – Citrullus lanatus

1gv1 – CvMDH

1b8p – AaMDH

7mdh – MDH – Sorgum bicolor

1mld – SsMDH

2cmd - EcMd+citrate

3nep – Md – Salinibacter ruber

Additional Resources
For additional information, see: Carbohydrate Metabolism